While RNA interference (RNAi) continues to hold incredible potential, numerous challenges associated with the application of RNAi technology must be addressed before it can be made into a viable therapy. The most prominent include transporting, targeting, and stabilizing short interfering RNAs (siRNAs) into tumor cells after injection into a patient's bloodstream. One of the most promising set of solutions to date includes the use of various types of nanoparticles (NPs) (Whitehead et al. 2009; Oh and Park 2009).
The rapidly expanding field of nanobiology opens up the possibilities for the development of new methods and compositions that can be used for the diagnosis, prognosis, and treatment of a multitude of diseases and conditions. However, while an increasing number of novel drugs and therapeutic agents are being discovered, the problem of delivering them specifically to the desired site or cell has not been solved. RNA nanoparticles have been shown to be able to carry multiple components, including molecules for specific cell recognition, image detection, and therapeutic treatment. The use of such protein-free nanoparticles holds the promise for the repeated long-term treatment of chronic diseases with low immune response and should avoid the problems of short retention time of small molecules and the difficulty of delivery of particles larger than 100 nanometers.
For example, NPs can provide several distinct advantages toward the advancement of RNAi therapeutics. For instance, they have been shown to produce a nanoparticle effect that improves cellular uptake. Moreover, NPs offer an increased degree of protection against ribonuclease degradation while also accommodating additional functional groups like aptamers to aid cellular targeting.
While a broad range of materials have been used in RNAi nanotechnology, including some exotic synthetic materials, unmodified RNA nucleotides that serve as both the therapeutic and the structural core of NPs are thought to provide unique advantages. For example, the use of natural RNA nucleotides—in addition to RNA's biocompatibility—takes advantage of RNA's inherent ability to self-assemble and spatially arrange multiple siRNAs, RNA or DNA aptamers, flourescent dyes, small molecules, RNA-DNA hybrids with split functionalities, and proteins. Furthermore, NPs made of unmodified nucleotides can be synthesized directly via run-off transcription, making their ease of synthesis and cost of production attractive for scaled-up production.
Formation of functional RNA NPs has been previously described and can take place either with one-pot assembly or directly with T7 RNA polymerase transcription reactions when equimolar amounts of DNA templates encoding specifically designed RNAs that are part of the composition of the functional RNA NPs (see, e.g. PCT/US2013/058492, incorporated by reference in its entirety herein).
Accordingly, there remains a need in the art for the development of siRNA nanoscaffolds to address several present challenges associated with NP-based siRNA delivery including cell-targeting, ease of synthesis, and triggered activation of therapeutic functionalities, and to provide a safe and efficient nanoparticle needs for the delivery of effective therapeutic and diagnostic siRNAs.